Antitoxin Levels Research Articles

Determining the Potency of Vaccines Containing Clostridium perfringens Epsilon Toxoid via Toxicity Analysis in MDCK Cell Lines

The potency control of the clostridium vaccines containing epsilon toxoid is performed with the Toxin Neutralization Test (TNT). Although TNT is a standard method, it has some disadvantages in terms of animal welfare and method validation studies. For this reason, the development and application of alternative methods are encouraged. However, purification and standardization of antigen, antibody and reference standards in serological-based in vitro methods still remain a significant problem. In this respect, cell culture analyses performed using similar reagents are considered possible alternatives in potency tests of the clostridial vaccines. Based on this approach, epsilon antitoxin levels in the sera of vaccinated rabbits were determined by cytotoxicity assay in mouse TNT and Madin-Darby Canine Kidney (MDCK) cell cultures. The toxicity response in cell culture is similar to the response in mice. A high correlation and a good linear relationship were observed in the results of the two methods. There were no non-specific reactions. These results show that, in potency tests of vaccines containing Clostridium perfringens epsilon toxoid, in vitro MDCK cell line seroneutralization assay can be successfully used instead of in vivo mouse TNT.

Primary diphtheria immunisation of adolescents and adults with low-dose vaccine, a survey of historic evidence from the literature.

The Public Health Agency of Sweden carried out a literature review on diphtheria vaccinations for seronegative people above 6 years of age with an uncertain vaccine history. The aim was to harmonise national Swedish recommendations with the current World Health Organization recommendations. There was no firm conclusion about dosage. Some low-dose vaccines used in the past had suboptimal potency, while others evoked adequate levels of antitoxin after three primary doses. We concluded that low-dose diphtheria vaccines that have been approved by a national medical products agency can be used for primary vaccination against diphtheria for individuals above 6 years of age.

Exposure-Response Modeling and Simulation to Support Human Dosing of Botulism Antitoxin Heptavalent Product.

Botulism antitoxin heptavalent (A, B, C, D, E, F, and G – Equine; BAT) product is a sterile solution of F(ab')2 and F(ab')2‐related antibody fragments prepared from plasma obtained from horses that have been immunized with a specific serotype of botulinum toxoid and toxin. BAT product is indicated for the treatment of symptomatic botulism following documented or suspected exposure to botulinum neurotoxin serotypes A to G in adults and pediatric patients. Pharmacokinetic and exposure‐response models were used to explore the relationship between BAT product exposure and the probability of survival, and the occurrence of relevant moderate clinical signs observed during the preclinical development of BAT product to justify the clinical dose. The predicted probability of survival in humans for all serotypes of botulinum neurotoxin was more than 95.9% following intravenous administration of one vial of BAT product. Furthermore, this BAT product dose is expected to result in significant protection against clinical signs in human adults for all botulinum neurotoxin serotypes. Our exposure response model indicates that we have sufficient antitoxin levels to give full protection at various theoretical exposure levels and, based on neutralization capacity/potency of one dose of BAT product, it is expected to exceed the amount of circulating botulinum neurotoxin.

Generalized Tetanus in a Case of an Immunized Patient

<h3>Purpose</h3> Tetanus is an infectious disease rarely occurring in developed countries, but its severity, and the possibility of developing multiple complications, some life-threatening, made its recognition and treatment to be a medical emergency. <h3>Methods & Materials</h3> The case report was based on the medical information from the database of County Emergency Clinical Hospital Oradea's (CECHO), Bihor region, Romania. <h3>Results</h3> The authors present the case of a 31-year-old female, with obesity, hyperinsulinemia, insulin resistance, intramuscular injections performed by unauthorized persons at the patient's home with 3 weeks before, and with a complete immunization antitetanic history, serum antitoxin level of 2.21 IU/mL, who presented to the emergency department of CECHO, in May 2021, for sore throat, trismus, opisthotonos induced by minimal external stimuli. C-reactive protein was slightly increased (35.9 mg/L), computed tomography and magnetic resonance imaging of the head showed no pathological changes. The diagnosis was based on the presence of clinical features. The patient followed specific etiological treatment. Metronidazole and Penicillin G were added as an antibiotic regimen. The patient responded slowly to the human tetanus immunoglobulin (5500 IU), in addition with one dose of tetanus toxoid adsorbed, with decreasing frequency and intensity of opisthotonos. <h3>Conclusion</h3> There are only a few reported cases of tetanus in immunized individuals in the medical literature (1). The mechanism of this is unclear. The antigenic variability, or an immune suppression, possibly mediated by leptin in the context of obesity, insulin resistance must be considered (1,2,3). The case emphasizes that the diagnosis of tetanus should not be rejected solely based on the presence of specific antibodies.

Should Tetanus Vaccine Booster be Given Earlier in Individuals Living with HIV?

The incidence rate of tetanus has dramatically decreased following the discovery of the tetanus vaccine. A decennial booster dose is necessary to maintain the protective antibody levels after the primary vaccination schedule. The recommendations for the tetanus booster doses in adult "people living with acquired immune deficiency virus (HIV)" (PLWH) is similar to those for the general population. However, the duration of protective antibodies in PLWH is unknown. The aim of this study was to determine the factors affecting the response of HIV-infected individuals to tetanus vaccine and to evaluate the role of tetanus antitoxin level in determining the timing of the booster dose. PLWH attending the Adult Vaccination Unit of Ege University Faculty of Medicine Infectious Diseases and Clinical Microbiology Department were tested for tetanus antibodies from 30 October to 30 November 2020. Demographic information and the history of primary vaccination and booster doses were derived from medical files. Tetanus antibodies were detected with "enzyme immunoassay (EIA)" method with Clostridium tetani toxin 5S IgG-"enzyme linked immunosorbent assay (ELISA)" kit (Nova Lisa, Novatec Immundiagnostica, Germany). Antibody levels <0.01 IU/ml were considered negative, 0.01-0.5 IU/ml weak positive, 0.51-1.0 IU/ml positive and > 1.1 IU/ml strong positive. A level ≥ 0.50 IU/ml was considered as protective. The study included 146 PLWH [men (n= 126) and women (n= 40)]. The mean age was 39.5 ± 11.20 years (range: 18-65). Protective antibody level was detected in 114 (78.1%) participants. Receiver operating characteristics (ROC) analysis revealed that the ideal lower limit of CD4+ T cell count during booster vaccination for a person to develop protective antibody level was calculated as ≥ 218 cell/mm3. In the multivariate analysis, it was found that age <50 years (OR= 16.4, 95% CI= 4.9-55.2), the absence of AIDS at the time of diagnosis (OR= 6.7, 95% Cl= 1.05-17.4) and CD4+ T cell count ≥ 218 cells/mm3 at the time of vaccination (OR= 4.2, 95% CI= 1.05-17.4) were associated with protective levels of antibody. Protective levels of tetanus antibodies in PLWH are lower than the general population. It should be considered that the booster dose may be required earlier, especially in PLWH who are > 50 years old, with CD4+ T lymphocyte count <200 cells/mm3 during previous booster vaccination and who had AIDS at the time of the diagnosis.

Insights into the Neutralization and DNA Binding of Toxin-Antitoxin System ParESO-CopASO by Structure-Function Studies.

ParESO-CopASO is a new type II toxin–antitoxin (TA) system in prophage CP4So that plays an essential role in circular CP4So maintenance after the excision in Shewanella oneidensis. The toxin ParESO severely inhibits cell growth, while CopASO functions as an antitoxin to neutralize ParESO toxicity through direct interactions. However, the molecular mechanism of the neutralization and autoregulation of the TA operon transcription remains elusive. In this study, we determined the crystal structure of a ParESO-CopASO complex that adopted an open V-shaped heterotetramer with the organization of ParESO-(CopASO)2-ParESO. The structure showed that upon ParESO binding, the intrinsically disordered C-terminal domain of CopASO was induced to fold into a partially ordered conformation that bound into a positively charged and hydrophobic groove of ParESO. Thermodynamics analysis showed the DNA-binding affinity of CopASO was remarkably higher than that of the purified TA complex, accompanied by the enthalpy change reversion from an exothermic reaction to an endothermic reaction. These results suggested ParESO acts as a de-repressor of the TA operon transcription at the toxin:antitoxin level of 1:1. Site-directed mutagenesis of ParESO identified His91 as the essential residue for its toxicity by cell toxicity assays. Our structure-function studies therefore elucidated the transcriptional regulation mechanism of the ParESO-CopASO pair, and may help to understand the regulation of CP4So maintenance in S. oneidensis.

Humoral Immune Response to Clostridioides difficile Toxins A and B in Hospitalized Immunocompromised Patients With C difficile Infection.

BackgroundThe humoral immune response to Clostridioides difficile toxins in C difficile infection (CDI) is incompletely characterized in immunocompromised hosts (ICHs).MethodsWe conducted a prospective study of hospitalized adults with CDI, with and without immunosuppression (hematologic malignancy, active solid tumor, solid organ or stem cell transplant, inflammatory bowel disease, autoimmune disease, congenital or acquired immunodeficiency, asplenia, chronic receipt of high-dose steroids, or receipt of immunosuppressing medications within 12 months). Serum and stool antibody concentrations of immunoglobulin (Ig)M, IgG, and IgA to C difficile toxins A and B at treatment days 0, 3, and 10–14 were compared.ResultsNinety-eight subjects (47 ICH; 51 non-ICH) were enrolled. Baseline serum antitoxin A and B antibody levels were similar. At day 3, ICHs demonstrated lower serum levels of antitoxin A IgG, antitoxin A IgA, and antitoxin B IgA (all P < .05). At day 10–14, lower antitoxin A IgG concentrations were observed in ICHs (ICH, 21 enzyme-linked immunosorbent assay [ELISA] units; interquartile range [IQR], 16.4–44.6) compared with non-ICH subjects (49.0 ELISA units; IQR, 21.5–103; P = .045). In stool, we observed lower concentrations of antitoxin B IgA antibodies at baseline and at day 3 for ICH subjects, with a notable difference in concentrations of antitoxin B IgA at day 3 (ICH, 6.7 ELISA units [IQR, 1.9–13.9] compared with non-ICH, 18.1 ELISA units [IQR, 4.9–31.7]; P = .003).ConclusionsThe ICHs with CDI demonstrated lower levels of C difficile antitoxin antibodies in serum and stool during early CDI therapy compared with non-ICHs. These data provide insight into the humoral response to CDI in ICHs.

Anti-toxin antibody is not associated with recurrent Clostridium difficile infection

Clostridium difficile infection (CDI) recurs in ∼20% of patients. Prior studies indicated that antibody responses directed against the C. difficile toxins A and B were potentially associated with lower risk of recurrent CDI. Here we tested the hypothesis that circulating anti-toxin IgG antibody levels associate with reduced risk of recurrent CDI. A cohort study with prospective enrollment and retrospective data abstraction examined antibody levels in 275 adult patients at the University of Michigan with CDI. We developed an enzyme linked immunosorbent assay to detect IgG antibodies against toxin A and toxin B in sera obtained at the time of diagnosis. Logistic regression examined the relationship between antibody levels and recurrence, and sensitivity tests evaluated for follow-up and survivor biases, history of CDI, and PCR ribotype. Follow-up data were available for 174 subjects, of whom 36 (20.7%) had recurrence. Comparing antibody levels vs. recurrence and CDI history, anti-toxin A levels were similar, while anti-toxin B levels had a greater range of values. In unadjusted analysis, detection of anti-toxin A antibodies, but not anti-toxin B antibodies, associated with an increased risk of recurrence (OR 2.71 [1.06, 8.37], P = .053). Adjusting for confounders weakened this association. The results were the same in sensitivity analyses. We observed a borderline increased risk of recurrence in patients positive for anti-toxin A antibodies, and sensitivity analyses showed this was not simply a reflection of prior exposure status. Future studies are needed to assess how neutralizing antibody or levels after treatment associate with recurrence.

YjbH Solubility Controls Spx in Staphylococcus aureus: Implication for MazEF Toxin-Antitoxin System Regulation.

Bacterial cells respond to environmental stresses by modulating their gene expression and adjusting their proteome. In Staphylococcus aureus, selective degradation by ClpP protease eliminates damaged proteins and regulates the abundance of functional proteins such as many important stress-induced transcriptional regulators. Degradation by ClpP requires the unfolding activity of partner Clp ATPases, such as ClpX and ClpC, and assistance of substrate-specific adaptor proteins such as YjbH and TrfA. Herein, we demonstrated that YjbH is aggregated in response to growth stress stimuli, such as oxidative and antibiotic stresses. In consequence, its function as an adaptor protein is compromised. YjbH controls the degradation of the stress-induced transcriptional regulator, Spx. Aggregated YjbH cannot assist Spx degradation, which results in Spx accumulation. We discovered that depending on the stress stimulus, Spx can be soluble or insoluble, and, consequently, transcriptionally active or inactive. Therefore, Spx accumulation and solubility are key components governing activation of Spx-dependent genes. Spx positively regulates expression of a ClpCP adaptor protein TrfA. TrfA in turn is required for degradation of MazE antitoxin, the unstable component of the MazEF toxin-antitoxin system, that neutralizes the endoribonuclease activity of MazF toxin. Bacterial toxin-antitoxin systems are associated with dormancy and tolerance to antibiotics that are related to chronic and relapsing infections, and it is at present a key unresolved problem in medicine. MazF activity was linked to growth stasis, yet the precise environmental signals that trigger MazE degradation and MazF activation are poorly understood. Here we propose a model where YjbH serves as a sensor of environmental stresses for downstream regulation of MazEF activity. YjbH aggregation, soluble Spx, and TrfA, coordinately control MazE antitoxin levels and consequently MazF toxin activity. This model implies that certain stress conditions culminate in modulation of MazF activity resulting in growth stasis during in vivo infections.

Seroprotection rate in adults after 1-dose, 2-dose, and 3-dose series of a reduced-diphtheria-tetanus toxoid vaccine (Td) during a diphtheria outbreak in Thailand

To evaluate seroprotection of different dosing strategies of reduced-diphtheria-tetanus-toxoid vaccine (Td) for adults during a diphtheria outbreak in Thailand, we enrolled 160 healthcare workers and 161 adults aged 20–60 years old and measured diphtheria antitoxin (DAT) level before administration of a Td vaccine. We scheduled a second Td at 4–8 weeks and a third Td at 6–12 months interval. DAT was measured 4 weeks after each dose. DAT levels of ≥0.1 and ≥1 IU/mL were considered as seroprotective and long-term seroprotective. Persons achieving long-term seroprotection were not given a further dose.The baseline seroprotection rate was 32.6%, which increased to 87.1% (95% confidence interval, 83.4–90.8%) after one dose. The seroprotection rate increased slightly with additional doses. The immune response was lowest among persons 30–49 years of age. We suggest 1-dose Td for adults during a diphtheria outbreak, and a 2-dose series being considered for those born before 1980.

Production, immunogenicity, stability, and safety of a vaccine against Clostridium perfringens beta toxins.

Background and Aim:The beta toxin is causing the most severe Clostridium perfringens-related diseases. This work was dedicated to developing a vaccine against beta toxin using C. perfringens type C (NCTC 3180).Materials and Methods:The crude toxoid harvest contained 710 limits of flocculation (Lf)/mL. The vaccine was formulated. Each 1 mL of the final vaccine product contained at least 50 Lf/mL of beta toxoids, 0.2 mL 3% aluminum hydroxide gel (equivalent to 5.18 mg of aluminum), <0.001% W/V thiomersal, formaldehyde <0.05% W/V, and ~0.7 mL phosphate-buffered saline (pH 7.2). The efficacy of the vaccine was evaluated by potency, stability, and safety tests.Results:The vaccine demonstrated 24.36 IU/mL (standard deviation, ±0.56) and 14.74 IU/mL (±0.36) of neutralizing antibodies in rabbits and cattle, respectively. Indeed, these levels were above the minimum recommended by international protocols since the obtained antibody levels had 2.43- and 1.47-fold increase in both rabbits and cattle, respectively, over the minimum antitoxin level suggested by the United States Department of Agriculture. Interestingly, our formulation was capable of inducing 1.65-fold higher immune responses in rabbits than that stimulated in cattle (65% increase) with a significant difference (p<0.0001). The vaccine was stable up to 30 months. The vaccinated rabbits were suffered from a temporarily slight increase in temperatures in the first 10 h without any significant difference (p>0.05).Conclusion:The research showed a procedure for the manufacturing process of the vaccine against C. perfringens beta toxins with a feasible quantity and the vaccine described here showed to be effective in eliciting levels of neutralizing antibodies higher than required by international standards. In addition, The vaccine was stable up to 30 months. Thus, it may represent an effective and safe for preventing C. perfringens-related diseases in rabbits and cattle, although further studies to prove its efficacy in the field on other farm animals are still needed.

Functionality and cross-regulation of the four SprG/SprF type I toxin-antitoxin systems in Staphylococcus aureus.

Toxin–antitoxin (TA) systems are ubiquitous among bacteria, frequently expressed in multiple copies, and important for functions such as antibiotic resistance and persistence. Type I TA systems are composed of a stable toxic peptide whose expression is repressed by an unstable RNA antitoxin. Here, we investigated the functionalities, regulation, and possible cross-talk between three core genome copies of the pathogenicity island-encoded ‘sprG1/sprF1’ type I TA system in the human pathogen Staphylococcus aureus. Except for SprG4, all RNA from these pairs, sprG2/sprF2, sprG3/sprF3, sprG4/sprF4, are expressed in the HG003 strain. SprG2 and SprG3 RNAs encode toxic peptides whose overexpression triggers bacteriostasis, which is counteracted at the RNA level by the overexpression of SprF2 and SprF3 antitoxins. Complex formation between each toxin and its cognate antitoxin involves their overlapping 3′ ends, and each SprF antitoxin specifically neutralizes the toxicity of its cognate SprG toxin without cross-talk. However, overexpression studies suggest cross-regulations occur at the RNA level between the SprG/SprF TA systems during growth. When subjected to H2O2-induced oxidative stress, almost all antitoxin levels dropped, while only SprG1 and SprF1 were reduced during phagocytosis-induced oxidative stress. SprG1, SprF1, SprF2, SprG3 and SprF3 levels also decrease during hyperosmotic stress. This suggests that novel SprG/SprF TA systems are involved in S. aureus persistence.

Diphtheria Carriage

The diphtheria carriage is a asympthomatic colonization of oro- and nasopharynx by toxigenic and nontoxigenic Corynebacterium diphtheriae. The carriage of toxigenic strains prevents a complete eradication of diphtheria infection in spite of mass toxoid immunization. The contamination by toxigenic diphtheria bacteria leads to the carriage if the person has a protective level of diphtheria antitoxin. Contamination with the toxigenic and nontoxigenic leads to the carriage if the person has no protection to the bacterial colonization factors. Some of them are surface protein structures and may serve as components of the future vaccines against diphtheria bacteria colonization.

Population Immunity to Diphtheria and Tetanus in the Republic of Belarus Following Long-Standing Vaccination

Study results of IgG to diphtheria and tetanus in 785 residents aged from 1 to 76 years old from different regions of the Republic of Belarus (in 2017) in long-term (since 1996) immunization schedule: at 3, 4, 5 and at 18 months old – DTP vaccine, at 6 years old – DT, at 11 years old – Diphtheria toxoid, at 16 years old, 26 years old and every following 10 years –Td or Diphtheria toxoid are presented. The antibody concentration was measured by Virion/Serion kits (Germany) and evaluated in accordance with the international standard: less than 0.01 IU/ml – individual is susceptible, 0.01–0.09 IU/ml – levels of antitoxin giving some degree of protection, 0.1 – &lt; 1 IU/ml – protective level of circulating antitoxin, ≥ 1.0 IU/ml – a level of antitoxin giving long-term protection. It was shown that the proportion of immune individuals against diphtheria and tetanus (with antibodies ≥ 0.01 IU/ml) was 96.7% (CI 95.4 ÷ 97.9) and 99.5% (CI 99.0 ÷ 100,0), respectively, and was quite high in all the population age groups – from 87.7 to 100% for diphtheria and from 96.5 to 100% for tetanus. In seropositive individuals IgG were presented mainly in protective and highly protective (≥ 0.1 IU/ml) titers: for diphtheria 93.7% – in 1–14 years old; 88.7% – in 15–19 years old; 78.4% – in 20–76 years old and for tetanus 100.0% – in 1–14 years old; 100.0% – in 15–19 years old; 99.3% – in 20–76 years old. Comparison of the current and previous studies results (in 1989–1994 – during the outbreak of diphtheria, in 1998–2001 – after the mass immunization campaign, in 2004 – in the context of continuous single cases of diphtheria registration in adults) had shown that the data of 2017 demonstrated the highest population immunity level to diphtheria and to tetanus in the last 30 years of observation.

Relationship Between Tetanus Antitoxin Titration Level and Vaccination History

Objectives: We aimed to determine tetanus antitoxin levels and to evaluate their relationship with history of vaccination among patients applying to the outpatient clinics of a University hospital. Methods: A questionnaire including socio-demographic characteristics and tetanus vaccination status was applied and blood samples taken from 218 subjects between 1 and 30 June 2015. Participants were classified into five groups according to their vaccination timing. Results: The mean age of participants was 46.7±15.4 years and 134 (61.5%) were women. Tetanus antitoxin levels were found weak positive in 54 (24.8%) patients, positive in 44 (20.2%) and strong positive in 120 (55.0%). Tetanus antitoxin level positivity was significantly associated with vaccination timing according to history. Among 105 participants who did not remember being vaccinated or who knew they were vaccinated but did not remember the date, 16 (15.2%) remembered the vaccination time when their injury, military service and pregnancy were questioned specifically. Antitoxin levels decreased with increasing age independent of gender (0.9-fold increase/year). Conclusion: We found that the booster dose recommended every 10 years was not applied sufficiently. Tetanus vaccination history must be questioned in more detail among people who do not remember/know their vaccination history, with specific questions regarding pregnancy, military service and injury histories.

Screening of Corynebacterium diphtheriae, Corynebacterium ulcerans and Corynebacterium pseudotuberculosis in throat swab specimens of children with upper respiratory tract infections

Although a significant decrease has been reported in the incidence of diphteria in many regions of the world following the routine diphtheria immunization programs, the emergence of new cases indicated that toxigenic strains are still circulating in the community. Diphtheria vaccine does not provide protection against asymptomatic carriage and colonization of non-toxigenic Corynebacterium diphtheriae. It is a known fact that invasive infections may arise from non-toxigenic C.diphtheriae strains that the non-toxigenic strains can become toxigenic strains leading to diphteria. It is also known that there is a risk of diphteria outbreaks due to decreased antitoxin level and inadequate adult immunization programs. In our country, there is no routine surveillance of toxigenic and non-toxigenic C.diphtheriae. In the present study we aimed to investigate the presence of C.diphtheriae, Corynebacterium ulcerans and Corynebacterium pseudotuberculosis in children presenting with the symptoms of upper respiratory tract infections that might be confused with those moderate diphteria, in order to highlight the requirement of microbiological surveillance and to create awareness about these microorganisms among public health experts, microbiologists and clinicians. Throat swab specimens were obtained from children who were admitted to the pediatric outpatient clinics, in Dr. Sami Ulus Obstectrics, Children Health and Diseases Educational and Research Hospital, with upper respiratory tract infections between 1 February 2016-22 March 2016. The specimens were inoculated in 5% sheep blood agar plates. The plates that were incubated in appropriate conditions, were evaluated for Group A beta hemolytic streptococcocci. Subsequently, culture plates were sent to the Public Health Institution of Turkey, National Respiratory Pathologens Reference Laboratories for the investigation of the presence of C.diphtheriae, C.ulcerans and C.pseudotuberculosis. The growth in each plate were collected with a sterile swab and inoculated in tryptic soy broth. Following 2 hours of incubation at 37oC, subcultures were inoculated in cystine-tellurite-blood agar (CTBA) and 5% sheep-blood agar plates; after an overnight incubation tellurite-reducing colonies were inoculated in Tinsdale agar plates. The suspected colonies with positive cystinase activity were identified by conventional methods and also with Coryne API (Biomerieux, France) systems. Toxicity tests (ELEK, PCR) were performed to investigate whether the C.diphtheriae strains were producing toxins. A total of 500 patients were involved in the study. Of these 260 (52%) were girls and 240 (48%) were boys with a mean age of 76 (range, 21-213) months. All patients except one were fully vaccinated with boosters. Most common presenting symptoms of the patients were fever (19.8%), sore throat (52.6%), cough (49.2%), tonsillar hyperemia (97.6%), presence of crypt (24.6%), and membrane over tonsils (1%). Group A beta-hemolytic streptococcocci were detected in the throat swab cultures of 66 (%13.2) patients. Genotypically toxin negative C.diphtheriae biovar gravis was identified in the throat swab cultures of 3 patients (2 girls and 1 boy). The tonsils were hyperemic and hypertrophic in all the patients with C.diphtheriae biovar gravis. C.ulcerans and C.pseudotuberculosis were detected in none of the patients. It is considered that similar regular cross-sectional studies or routine screening programs are expected to raise awareness about this forgotten microorganism both epidemiologically and microbiologically.

Estimation of Time Period for Effective Human Inhalational Anthrax Treatment Including Antitoxin Therapy.

Introduction:Infrequent natural human inhalational anthrax cases coupled with high bioterrorism risk have brought about use of animal models to serve as the basis for approval of novel treatments. For inhalational anthrax, protective antigen (PA) drives much of the mortality, and raxibacumab, an anti-PA monoclonal antibody, has been approved for therapeutic use using the Animal Rule. Given the paucity of human inhalational anthrax clinical data including PA kinetics, the post-exposure period for effective treatment of human disease remains unknown. The objective of this investigation was to extrapolate animal PA kinetics to a conceptual human model to estimate the post-exposure period for effective treatment of human inhalational anthrax.Methods:Human PA kinetic parameters were extrapolated from reported rabbit and monkey data. PA profiles were simulated with and without antibiotic induced PA clearance to represent antibiotic-sensitive and -resistant infections, respectively. Antitoxin levels equimolar to or greater than concurrent PA levels were considered protective.Results:For antibiotic sensitive infections, treatment with antibiotics alone ≤4 days after spore exposure prevents toxemia. Administration of raxibacumab together with antibiotics protects ≥ 80% of subjects for 3 additional days (7 days post exposure). In the setting of antibiotic resistance, raxibacumab would be protective for at least 6 days post exposure.Conclusions:Although the animal model of disease does not reflect the potential impact of supportive care (e.g. fluid resuscitation received by critically ill patients) on PA kinetics and raxibacumab PK, the simulations suggest that administration of antitoxin in combination with antibiotics should provide a longer postexposure window for effective treatment than for antibiotics alone. In addition, raxibacumab administration soon after exposure to an antibiotic resistant strain should provide effective treatment.

Immunity to Diphtheria and Tetanus in Army Personnel and Adult Civilians in Mashhad, Iran.

This study aimed to investigate serologic immunity to diphtheria and tetanus in army personnel and a sample population of adult civilians in Mashhad, Iran. Army personnel (n = 180) and civilians (n = 83) who presented at Mashhad army hospital participated in this study. Diphtheria and tetanus antitoxin levels were determined by enzyme-linked immunosorbent assay. Approximately 77% and 94% of army personnel aged 18-34 years had at least basic protection against diphtheria (antitoxin level ≥0.1 IU/mL) and tetanus (antitoxin level >0.1 IU/mL), respectively. For civilians in this age group, the proportions were 76% for both diseases. Antitoxin levels waned with age. Thus, participants older than 50 years had lower immunity; this decrease in immunity was more pronounced for tetanus than for diphtheria in both army personnel and civilians. For both diseases, geometric mean antitoxin titers and the proportion of participants with at least basic protection were higher in subjects with a history of vaccination in the last 10 years (P < 0.001), higher in men than women, and in army personnel than civilians in each age group. Young army personnel and civilians (18-34 years old) had adequate immunity to diphtheria and tetanus. However, the large number of susceptible older adults (>50 years old) calls for improved booster vaccination protocols.

Long-Term Protection against Diphtheria in the Netherlands after 50 Years of Vaccination: Results from a Seroepidemiological Study.

Background and AimsTo evaluate the National Immunisation Programme (NIP) a population-based cross-sectional seroepidemiological study was performed in the Netherlands. We assessed diphtheria antitoxin levels in the general Dutch population and in low vaccination coverage (LVC) areas where a relatively high proportion of orthodox Protestants live who decline vaccination based on religious grounds. Results were compared with a nationwide seroepidemiological study performed 11 years earlier.MethodsIn 2006/2007 a national serum bank was established. Blood samples were tested for diphtheria antitoxin IgG concentrations using a multiplex immunoassay for 6383 participants from the national sample (NS) and 1518 participants from LVC municipalities. A cut-off above 0.01 international units per ml (IU/ml) was used as minimum protective level.ResultsIn the NS 91% of the population had antibody levels above 0.01 IU/ml compared to 88% in the 1995/1996 serosurvey (p<0.05). On average, 82% (vs. 78% in the 1995/1996 serosurvey, p<0.05) of individuals from the NS born before introduction of diphtheria vaccination in the NIP and 46% (vs. 37% in the 1995/1996 serosurvey, p = 0.11) of orthodox Protestants living in LVC areas had antibody levels above 0.01 IU/ml. Linear regression analysis among fully immunized individuals (six vaccinations) without evidence of revaccination indicated a continuous decline in antibodies in both serosurveys, but geometric mean antibodies remained well above 0.01 IU/ml in all age groups.ConclusionsThe NIP provides long-term protection against diphtheria, although antibody levels decline after vaccination. As a result of natural waning immunity, a substantial proportion of individuals born before introduction of diphtheria vaccination in the NIP lack adequate levels of diphtheria antibodies. Susceptibility due to lack of vaccination is highest among strictly orthodox Protestants. The potential risk of spread of diphtheria within the geographically clustered orthodox Protestant community after introduction in the Netherlands has not disappeared, despite national long-term high vaccination coverage.

Seroepidemiological Studies on Pertussis, Diphtheria and Tetanus among Nursing Students Enrolled between 1994 and 2011 in the Kitakyushu District

Seroepidemiological studies on pertussis, diphtheria and tetanus were performed on 1,540 nursing students enrolled in S-university between 1994 and 2011. Antibody titers against pertussis toxin (PT) and filamentous hemagglutinin (FHA), diphtheria antitoxin titer, and tetanus antitoxin titer were measured using sera taken during enrollment. The antibody-seropositive rates and geometric mean titers (GMTs) were calculated by according to birth year (1975-1993). The pertussis anti-PT and anti-FHA antibody-seropositive rates (seropositive levels were both defined as ≥10 EU/mL) were 12%-53% and 47%-84%, respectively. The anti-PT antibody-seropositive rate was lower than the anti-FHA rate. The anti-PT antibody GMTs were 2.0-11 EU/mL, whereas the anti-FHA antibody GMTs were 10 EU/mL or more (8.8-31 EU/mL) in almost every group. The diphtheria antitoxin-seropositive rate (≥0.1 IU/mL) was 49%-79%, and the GMT was nearly 0.1 IU/mL. The tetanus antitoxin-seropositive rate (≥0.01 IU/mL) was 91%-100%, and the GMT was 0.3 IU/mL or more for all the groups. While the nursing students' diphtheria and tetanus antitoxin levels were sufficient to prevent both diseases, the anti-PT antibody-seropositive rate and the GMT were both lower than the level required to prevent pertussis, suggesting that many of the nursing students were pertussis-susceptible. These findings suggest a need for pertussis vaccination in young adults or early adolescents.